肝孤核受体激动剂对间充质干细胞的抗缺氧复氧损伤作用及机制

目的探讨肝孤核受体(LXR)激动剂对脂肪间充质干细胞(AD-MSCs)缺氧损伤的保护作用及可能机制。方法酶消化法分离稳定表达萤火虫荧光素酶(Fluc)报告基因的小鼠AD-MSCs,流式细胞术检测CD90、CD44、CD34、CD45细胞表面标记物。3代AD-MSCs分为7组:对照组;缺氧6 h/复氧2 h组(缺氧复氧组);缺氧/复氧+DMSO组(DMSO组);缺氧复氧+不同浓度LXR激动剂T0901317干预组(1μmol/L组、5μmol/L组、10μmol/L组、15μmol/L组)。运用Fluc报告基因生物发光成像技术对AD-MSCs细胞增殖进行定量评价,免疫印迹法检测细胞NF-κB表达水平,ELISA法检测AD-MSCs的白细胞介素6(IL-6)、TNF-α分泌水平。结果流式细胞术结果显示.AD-MSCs呈CD44(+)、CD90(+)、CD34(-)、CD45(-)。光学成像结果显示,AD-MSCs细胞数与Fluc平均生物发光信号强度呈正相关(r~2=0.97)。与对照组比较,缺氧复氧组AD-MSCs分泌TNF-α、IL-6、NF-κB明显升高;与缺氧复氧组比较,10μmol/L组AD-MSCs分泌TNF-α、IL-6、NF-κB明显减少(P<0.05,P<0.01)。结论 LXR激动剂可以促进缺氧复氧损伤后AD-MSCs的存活,并能通过NF-κB信号途径抑制炎性因子IL-6、TNF-α的释放,可为干细胞移植治疗心肌缺血再灌注损伤提供新的细胞保护方法。     

白藜芦醇抑制缺氧复氧诱导心肌微血管内皮细胞凋亡的研究

目的探讨白藜芦醇对缺氧复氧诱导心肌微血管内皮细胞(CMEC)凋亡的抑制作用及其可能的分子机制。方法体外培养大鼠CMEC,建立缺氧复氧损伤模型,随机分为对照1组、缺氧复氧1组、缺氧复氧+白藜芦醇组(白藜芦醇1组),白藜芦醇分别选择5、10、20、30及40μmol/L浓度,MTT法检测CMEC增殖能力。20μmol/L白藜芦醇为最适浓度,使用磷脂酰肌醇3激酶(PI3K)特异性抑制剂LY294002,后续实验又分为对照2组、缺氧复氧2组、缺氧复氧+白藜芦醇2组(白藜芦醇2组)、缺氧复氧+白藜芦醇+LY294002组(LY294002组)。PI-AnnexinⅤ检测CMEC的凋亡;Western blot法检测细胞蛋白激酶B(Akt)、磷酸化Akt、缺氧诱导因子1α(HIF-1α)蛋白表达或磷酸化水平。结果与缺氧复氧1组比较,不同浓度白藜芦醇1组均可增加CMEC增殖能力,呈剂量依赖性,以白藜芦醇1组20μmol/L保护作用最显著(P<0.05)。与白藜芦醇2组比较,LY294002组CMEC凋亡率明显升高,磷酸化Akt和Akt蛋白、HIF-1α蛋白表达明显下降(P<0.05)。结论白藜芦醇可显著抑制缺氧复氧诱导的CMEC凋亡,其机制可能与PI3K/Akt介导的HIF-1α上调相关。     

法舒地尔后适应对大鼠心肌缺血再灌注损伤的保护机制研究

目的通过主动脉根部模拟冠状动脉内给药,探讨盐酸法舒地尔后适应对大鼠急性心肌缺血再灌注损伤的保护机制。方法选择SD大鼠30只,随机分为假手术组、缺血再灌注组、缺血后适应组、法舒地尔组、法舒地尔+LY294002组(LY294002组),每组6只。各组于再灌注180min后处死大鼠,取心肌组织用Western blot方法测定Bcl-2、Bax、caspase-3、蛋白激酶B(Akt)及磷酸化Akt的表达。结果与缺血再灌注组比较,缺血后适应组和法舒地尔组Bcl-2表达明显升高,Bax、caspase-3表达明显降低(P<0.05)。与法舒地尔组比较,LY294002组Bcl-2表达明显降低,Bax、caspase-3表达明显升高(P<0.05)。与缺血再灌注组和LY294002组比较,法舒地尔组磷酸化Akt表达明显升高(P<0.05)。结论盐酸法舒地尔后适应的机制可能与激活磷脂酰肌醇3-激酶-Akt传导通路有关。     

蛋白激酶B信号通路的干预在慢性脑低灌注大鼠中的作用及对海马细胞凋亡的影响

目的探讨蛋白激酶B/糖原合成酶激酶3β(Akt/GSK3β)信号通路是否参与慢性脑低灌注(CCH)大鼠海马细胞凋亡过程。方法选择SD大鼠60只,随机分为假手术组、CCH组、LY294002组、胰岛素样生长因子1(IGF-1)组,每组15只,后3组制作大鼠CCH模型,LY294002组和IGF-1组分别以抑制剂LY294002及激动剂IGF-1进行干预。采用TUNEL检测海马细胞凋亡;Western blot法检测Akt、GSK3β、磷酸化Akt(P-Akt)和磷酸化GSK3β(P-GSK3β)表达。结果与假手术组比较,CCH组、LY294002组及IGF-1组细胞凋亡水平均升高(P<0.05);与CCH组比较,LY294002组细胞凋亡数明显增高(20.33±1.52 vs 15.00±1.00,P<0.05),而IGF-1组细胞凋亡数降低(12.00±1.73 vs 15.00±1.00,P<0.05)。与假手术比较,CCH组和IGF-1组P-Akt,P-GSK3β均明显增高(P<0.05),而LY294002组无明显变化(P>0.05);与CCH组比较,IGF-1组P-Akt,P-GSK3β表达量均升高(P<0.05);LY294002组均降低(P<0.05)。结论 Akt/GSK3β信号通路参与CCH大鼠海马细胞凋亡过程,上调该通路Akt与GSK3β这2种蛋白的磷酸化表达水平,可能是拮抗这种细胞凋亡方式之一。     

H_2S对外源性LY294002预处理人结肠癌细胞凋亡的影响

目的研究H_2S对体外培养人结肠癌细胞凋亡的影响及其机制。方法体外培养人结肠癌细胞,随机分为正常对照组、NaHS(100μmol/L)组、LY294002(2μg/ml)组、NaHS(100μmol/L)+LY294002(2μg/ml)组,采用流式细胞术检测各组干预48 h后对人结肠癌细胞凋亡的影响,Western印迹检测各组干预48 h后对人结肠癌细胞中磷酸化Akt、非磷酸化GSK-3β蛋白表达的影响。结果与正常对照组比较,NaHS组人结肠癌细胞凋亡显著降低,p-Akt蛋白表达显著增加,而GSK-3β蛋白表达显著降低(均P<0.01)。LY294002组人结肠癌细胞凋亡显著增加,p-Akt蛋白表达显著降低,而GSK-3β蛋白表达均显著增加(均P<0.01)。而与NaHS组比较,NaHS+LY294002组人结肠癌细胞凋亡显著增加,p-Akt蛋白表达显著降低,而GSK-3β蛋白表达显著增加(均P<0.01)。结论 PI3K/Akt/GSK-3β信号通路可能介导参与了H_2S对人结肠癌细胞凋亡的抑制作用。     

CXCL8/PI3K-Akt信号通路在缺氧诱导的人脑胶质瘤细胞血管生成拟态中的作用

目的探讨CXCL8/PI3K-Akt信号转导通路在缺氧诱导的人脑胶质瘤细胞株U87MG和U251血管生成拟态中的作用。方法用氯化钴(Co Cl2)模拟缺氧环境,将人脑胶质瘤细胞株U87MG和U251置中培养。分别加入PI3K-Akt信号通路激动剂CXCL8及PI3K-Akt信号通路抑制剂LY294002干预48 h,体外成管实验检测各组细胞成管能力;Western印迹检测各组Akt、P-Akt蛋白的表达情况。结果缺氧可导致U87MG细胞和U251细胞形成的管腔样结构明显增加,Akt蛋白的磷酸化活化表达显著增加(P0.05)。加入CXCL8干预可使管腔样结构直径进一步增长,而加入LY294002干预48 h则可使其管腔样结构在数量和直径上显著少于缺氧组(P0.05)。CXCL8能够有效激活Akt磷酸化,LY294002则显著抑制其磷酸化活化(P0.05),而总Akt蛋白表达无差异(P0.05)。结论 CXCL8/PI3K-Akt信号转导通路在缺氧诱导的人脑胶质瘤细胞株U87MG和U251血管生成拟态中发挥重要的促进作用,可望成为胶质瘤治疗有效靶点。     

磷酯酰肌醇3-激酶/蛋白激酶B信号通路介导葛根素改善缺氧复氧损伤的作用及机制

目的:探讨磷酯酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)信号通路介导葛根素改善缺氧复氧损伤的作用及机制。方法:应用噻唑蓝法筛选出合适的葛根素以及PI3K抑制剂LY294002干预人脐静脉内皮细胞EA.hy926的浓度。分为对照组、缺氧复氧组、缺氧复氧+葛根素组(葛根素组),缺氧复氧+葛根素+LY294002处理组(葛根素+抑制剂组),每组均设3个复孔。应用蛋白免疫印迹法(Western blot)检测葛根素预处理细胞后以及葛根素预处理细胞的同时加入LY294002后对B细胞淋巴瘤/白血病-(2Bcl-2)、Bcl-2相关X蛋白(Bax)、活化天冬氨酸蛋白水解酶-(3cleaved caspase-3)、活化天冬氨酸蛋白水解酶-9(cleaved caspase-9)、磷酸化的蛋白激酶B(p-Akt)、Akt蛋白表达的影响。应用实时荧光定量PCR检测葛根素预处理对EA.hy926细胞缺氧复氧后天冬氨酸蛋白水解酶-3(caspase-3)和Bcl-2信使核糖核酸(m RNA)表达的影响。应用Hoechest染色检测葛根素预处理对EA.hy926细胞凋亡的影响。结果:葛根素(10-7mol/L)明显升高缺氧复氧后EA.hy926细胞活力(P0.001),并显著上调Bcl-2蛋白水平的表达,下调Bax、cleaved caspase-9、cleaved caspase-3蛋白水平的表达(P均0.01)。与缺氧复氧组相比,葛根素预处理后,p-Akt蛋白表达水平明显升高(P0.01)。加入LY294002(10μmol/L)后p-Akt蛋白表达水平下降,且抗凋亡蛋白Bcl-2的表达水平下降,促凋亡蛋白cleaved caspase-3、cleaved caspase-9、Bax的表达水平升高(P均0.01)。结论:葛根素对缺氧复氧诱导的EA.hy926细胞凋亡具有抑制作用,其抑制作用可能与其对PI3K/Akt信号通路的调控有关。     

DIDS通过PI3-K/Akt途径减弱缺血/再灌注损伤诱导心肌细胞凋亡

目的 探讨磷脂酰肌醇3激酶(PI3-K)/蛋白激酶B(Akt)信号通路在DIDS(4,4′-diisothiocyanostilbene-2,2′-disulfonic acid)减弱缺血/再灌注损伤（I/RI）诱导心肌细胞凋亡中的作用。方法 以I/RI诱导心肌细胞凋亡,然后用PI3-K特异性的抑制剂LY294002［22(42吗啉基)282苯基24氢212苯并吡喃242酮］进行干预。实验分为正常对照组、I/RI组、I/RI+DIDS组和I/RI+DIDS+LY294002组。通过噻唑蓝(MTT)比色法、Hoechest-33258染色和半胱天冬蛋白酶（Apo-ONETM Homogeneous Caspase）-3试剂盒以及Western blot分别检测:心肌细胞的存活率（%）、细胞核的形态变化和caspase-3活性、Akt的磷酸化。结果 ①DIDS能够显著抑制I/RI诱导的细胞存活率的下降,抑制凋亡小体的出现,抑制caspase-3的活性的增加(P<0.01)。②用LY294002预处理后,DIDS保护I/RI心肌细胞存活的作用减弱、凋亡小体增加和caspase-3活性明显升高(P<0.01)。③I/RI组与正常对照组Akt磷酸化无明显差异,DIDS能够显著增加I/RI组中Akt蛋白的磷酸化(P<0.01)。用LY294002预处理后,DIDS对I/RI组Akt蛋白磷酸化的影响明显减弱(P<0.01)。结论 DIDS可通过激活PI3-K/Akt信号通路减弱I/RI 诱导的心肌细胞的凋亡。     

葛根素通过PI3K/Akt途径对大鼠缺血再灌注心肌细胞凋亡的抑制

目的 观察葛根素预处理对缺血再灌注(I/R)大鼠心肌细胞凋亡的影响及其与PI3K/Akt信号通路的关系.方法 30只SD大鼠随机分为假手术组(SH组)、缺血再灌注组(I/R组)、葛根素预处理组(PU组)、葛根素预处理+LY294002组(PU+ LY组)及缺血再灌注+LY294002组(I/R+ LY组).除假手术组外,其他各组大鼠均行结扎冠状动脉前降支30 min,再灌注120 min,监测血流动力学,Western印迹法检测心肌组织p-Akt信号蛋白的表达;TUNEL法检测心肌细胞凋亡.结果 与I/R组比较,PU组能够明显改善心功能,抑制心肌细胞凋亡,并增加Akt的磷酸化水平(P<0.05,P<0.01).结论 葛根素预处理能够抑制缺血再灌注所致的心肌细胞凋亡,其作用机制与PI3K/Akt信号通路活化有关.     

阻断PI3K/Akt通路增强缺氧环境胰腺癌细胞化疗敏感性的实验研究

目的研究LY294002联合吉西他滨是否能增强缺氧环境下胰腺癌细胞株PANC-1的化疗敏感性。方法缺氧条件下培养胰腺癌细胞株PANC-1,运用MTT检测LY294002联合吉西他滨对胰腺癌细胞株细胞PANC-1生长的影响,运用Western blot检测AKT及磷酸化AKT蛋白表达的水平。结果 MTT检测在缺氧环境下LY294002联合吉西他滨作用的胰腺癌细胞PANC-1与单纯吉西他滨作用组相比细胞生存率显著下降(P=0.003),且具有时间依赖性。Western blot检测显示在缺氧环境下,LY294002联合吉西他滨作用的胰腺癌细胞PANC-1磷酸化AKT蛋白水平与单纯吉西他滨作用组相比显著降低(P=0.002)。结论阻断PI3K/Akt通路能增强缺氧环境下胰腺癌细胞化疗敏感性,为胰腺癌的治疗方法及逆转耐药提供了新的实验依据。     

Apelin enhances cardiac neovascularization after myocardial infarction by recruiting aplnr+ circulating cells

Rationale: Neovascularization stimulated by local or recruited stem cells after ischemia is a key process that salvages damaged tissue and shows similarities with embryonic vascularization. Apelin receptor (Aplnr) and its endogenous ligand apelin play an important role in cardiovascular development. However, the role of apelin signaling in stem cell recruitment after ischemia is unknown. Objective: To investigate the role of apelin signaling in recruitment after ischemia. Methods and Results: Aplnr was specifically expressed in circulating cKit+/Flk1+ cells but not in circulating Sca1+/Flk1+ and Lin+ cells. cKit+/Flk1+/Aplnr+ cells increased significantly early after myocardial ischemia but not after hind limb ischemia, indicative of an important role for apelin/Aplnr in cell recruitment during the nascent biological repair response after myocardial damage. In line with this finding, apelin expression was upregulated in the infarcted myocardium. Injection of apelin into the ischemic myocardium resulted in accelerated and increased recruitment of cKit+/Flk1+/Aplnr+ cells to the heart. Recruited Aplnr+/cKit+/Flk1+ cells promoted neovascularization in the peri-infarct area by paracrine activity rather than active transdifferentiation, resulting into cardioprotection as indicated by diminished scar formation and improved residual cardiac function. Aplnr knockdown in the bone marrow resulted in aggravation of myocardial ischemia-associated damage, which could not be rescued by apelin. Conclusions: We conclude that apelin functions as a new and potent chemoattractant for circulating cKit+/Flk1+/Aplnr+ cells during early myocardial repair, providing myocardial protection against ischemic damage by improving neovascularization via paracine action.     

丝素蛋白/壳聚糖复合纳米纤维膜心脏补片的生物相容性研究

目的 探讨丝素蛋白(SF)/壳聚糖(CS)复合纳米纤维膜对小鼠脂肪间充质干细胞（AD-MSCs）黏附和生长的影响，评估该材料用作心脏补片的可行性。 方法 通过静电纺丝技术制备纤维素纳米纤维底板，用层层自组装技术(LBL)，将SF和CS组装到纤维素纳米纤维表面，得到改性的SF/CS复合纳米纤维膜，用ζ- 电位和X射线光电子能谱(XPS)测定材料的表征。分离培养携带绿色荧光蛋白（GFP）和荧光素酶（Fluc）双重报告基因的小鼠AD-MSCs细胞，并接种于SF/CS复合纳米纤维膜上共培养，用激光共聚焦显微镜(CFM)观察细胞黏附和生长形态，扫描电子显微镜(SEM)观察超微结构，生物发光成像(BLI)及MTT比色法评估AD-MSCs细胞的数量和活性。结果 XPS测定的结果提示SF/CS复合纳米纤维膜构建成功。SEM观察可见纳米纤维直径均一，具有良好的三维多孔结构，细胞在SF/CS复合纳米纤维膜上增殖活跃，形态和生长状态良好。CFM、BLI及MTT比色法的结果提示细胞与SF/CS复合纳米纤维膜共培养后呈三维分布，增殖状态好。 结论 对纤维素纳米纤维用LBL进行改性后可明星促进AD-MSCs的三维生长，SF/CS复合纳米纤维膜是可用于组织工程心肌补片的理想材料。     

匹他伐他汀促进小鼠血管新生作用及其机制的研究

目的探讨匹他伐他汀对小鼠血管新生的促进作用及其可能的作用机制。方法建立野生型C3H／He小鼠下肢缺血模型并分为2组,缺血对照组,匹他伐他汀组。使用激光多普勒血流测定仪测定实验小鼠投药前、下肢缺血手术后双下肢血流。免疫荧光组化sP法计数缺血肢毛细血管数。免疫酶组化直接法计数缺血肢磷酸化蛋白激酶Akt（p-Akt）阳性细胞数。蛋白印迹杂交方法检测缺血肢血管内皮生长因子蛋白表达。镉还原Griess法测定实验结束后血清一氧化氮代谢产物含量。结果匹他伐他汀使实验小鼠术后缺血肢血流恢复明显,缺血肢与非缺血肢血流面积比明显增加;缺血肢毛细血管密度明显增加、p-Akt活性增加（p-Akt阳性细胞数明显增加）;血中一氧化氮代谢产物含量明显增高;缺血肢血管内皮生长因子蛋白表达增强。结论匹他伐他汀有促进小鼠血管新生的作用。     

Transplantation of Cardiac Mesenchymal Stem Cell-Derived Exosomes for Angiogenesis

We demonstrated the effects of exosomes secreted by cardiac mesenchymal stem cells (C-MSC-Exo) in protecting acute ischemic myocardium from reperfusion injury. To investigate the effect of exosomes from C-MSC on angiogenesis, we injected C-MSC-Exo or PBS intramuscularly into ischemic hind limb. Blood perfusion of limb was evaluated by laser Doppler Imaging. We observed that ischemic limb treated with C-MSC-Exo exhibits improved blood perfusion compared to ischemic limb treated with PBS at 2 weeks and 1 month after induction of limb ischemia. To explore the potential mechanisms underlying C-MSC-Exo’s angiogenetic effect, we performed microRNA array analysis and identify mmu-miR-7116-5p as the most abundant enriched miRNA detected in C-MSC-Exo. Bioinformatics’ analysis shows that miR-7116-5p negatively regulates protein polyubiquitination. In conclusion, our study demonstrated that intramuscular delivery of C-MSC-Exo after limb ischemia improves blood perfusion, and we identified the most abundant miRNAs that are preferentially enriched in C-MSC-Exo.     

Neovascularization in a mouse model via stem cells derived from human fetal amniotic membranes

In this study, we evaluated the effect of culture-expanded mesenchymal stem cells (MSCs), derived from amniotic membranes, on neovascularization and blood flow, in an animal model of limb ischemia in immune-deficient mice. MSCs were cultured from human amniotic membranes by collagenase digestion. Human amniotic mesenchymal stem cells (hAMSCs) were administered intramuscularly at three different sites of the ischemic leg whose femoral vessels were ligated. After 4 weeks of culture, a population of homogeneous mesenchymal cells was isolated from the human amniotic membranes after confluence was reached. We performed three different groups of mice model [controls, hAMSCs, conditioned media from the hAMSCs (hAMSCs-CM)]. The blood flow recovery in the hindlimb ischemia model was significantly higher in the hAMSC-transplanted group than in the control group. Moreover, hAMSCs-CM significantly improved the cutaneous blood flow. The histological examination showed that red fluorescence (CM-DiI)-labeled hAMSCs was detected in the interstitial tissues between the muscle fibers 2 weeks after transplantation. The results of this study showed that hAMSCs may be an attractive, alternative source of progenitor or stem cells for basic research as well as clinical applications.     

Potentiation of biological effects of mesenchymal stem cells in ischemic conditions by melatonin via upregulation of cellular prion protein expression

Mesenchymal stem cells (MSCs) are promising candidates for stem cell‐based therapy in ischemic diseases. However, ischemic injury induces pathophysiological conditions, such as oxidative stress and inflammation, which diminish therapeutic efficacy of MSC‐based therapy by reducing survival and functionality of transplanted MSCs. To overcome this problem, we explored the effects of melatonin on the proliferation, resistance to oxidative stress, and immunomodulatory properties of MSCs. Treatment with melatonin enhanced MSC proliferation and self‐renewal via upregulation of cellular prion protein (PrP^C) expression. Melatonin diminished the extent of MSC apoptosis in oxidative stress conditions by regulating the levels of apoptosis‐associated proteins, such as BCL‐2, BAX, PARP‐1, and caspase‐3, in a PrP^C‐dependent manner. In addition, melatonin regulated the immunomodulatory effects of MSCs via the PrP^C‐IDO axis. In a murine hind‐limb ischemia model, melatonin‐stimulated MSCs improved the blood flow perfusion, limb salvage, and vessel regeneration by lowering the extent of apoptosis of affected local cells and transplanted MSCs as well as by reducing infiltration of macrophages. These melatonin‐mediated therapeutic effects were inhibited by silencing of PrP^C expression. Our findings for the first time indicate that melatonin promotes MSC functionality and enhances MSC‐mediated neovascularization in ischemic tissues through the upregulation of PrP^C expression. In conclusion, melatonin‐treated MSCs could provide a therapeutic strategy for vessel regeneration in ischemic disease, and the targeting of PrP^C levels may prove instrumental for MSC‐based therapies.     

低氧诱导因子1α抑制大鼠缺血下肢血管生成及内皮祖细胞的动员

目的探讨低氧诱导因子1α(HIF-1α)在组织缺血时血管生成及内皮祖细胞(EPCs)动员中的作用。方法SD雌性大鼠36只,随机分为下肢缺血加HIF-1α反义寡核苷酸组(HIF-1α反义寡核苷酸组)、下肢缺血加HIF-1α错义寡核苷酸组(HIF-1α错义寡核苷酸组)和单纯下肢缺血组,每组12只,于股动脉结扎建立下肢缺血模型,缺血局部微注射脂质体转染的全硫代修饰HIF-1α寡核苷酸。测血浆血管内皮生长因子(VEGF);acLDL-DiI和FITC-UEA-I双染色鉴定EPCs后计数;下肢缺血区肌肉组织测HIF-1αmRNA和VEGF mRNA的表达和术后28天缺血区肌肉组织毛细血管密度。结果HIF-1α反义寡核苷酸组的HIF-1αmRNA和VEGF mRNA表达均较单纯下肢缺血组和HIF-1α错义寡核苷酸组明显减少;HIF-1α反义寡核苷酸组的循环EPCs、血浆VEGF及缺血区肌肉组织毛细血管密度均较单纯下肢缺血组和HIF-1α错义寡核苷酸组明显减少。结论全硫代修饰HIF-1α反义寡核苷酸可抑制大鼠缺血肢体血管生成和因缺血引起的EPCs的动员,其机制可能是通过VEGF起作用,HIF-1α是组织缺血时EPCs一个动员因子。     

G-CSF and HGF: combination of vasculogenesis and angiogenesis synergistically improves recovery in murine hind limb ischemia

Granulocyte colony-stimulating factor (G-CSF) is known to mobilize bone marrow stem cells into the peripheral circulation. This study was designed to investigate whether G-CSF by itself or in combination with hepatocyte growth factor (HGF) can promote vasculogenesis and angiogenesis in murine hind limb ischemia. Hind limb ischemia was induced in BALB/c nude or C57/BL6 mice that received bone marrow transplantation from green fluorescent protein (GFP)-transgenic mice. In the HGF group, hHGF expression plasmid was injected into the ischemic muscles. In the G-CSF group, G-CSF was administered subcutaneously for 10 days. The G-CSF+HGF group was concomitantly treated with G-CSF and HGF, and the control group received no treatment. All effects were confirmed at 4 weeks. The G-CSF+HGF group had a higher laser Doppler blood perfusion index, higher microvessel density, and a lower incidence of hind limb necrosis than the other groups. Confocal laser microscopy revealed that a number of GFP-positive cells infiltrated to the vasculature of the ischemic area. Some of the GFP positive cells were clearly co-immunostained with alpha-smooth muscle actin as well as von Willebrand factor. G-CSF-mobilized stem cells co-expressed CD49d and CD34, which would have promoted their adhesion to cells in the ischemic muscle that expressed HGF-induced vascular cell adhesion molecule-1. The combination of G-CSF and HGF had a significant synergistic effect, suggesting that the combination of mobilization of stem cells from bone marrow to peripheral circulation and their recruitment to the ischemic area might potentiate angiogenesis and vasculogenesis.     

超声心动图评估肺高血压的应用进展

心脏瓣膜钙化是由多种分子机制共同参与的复杂病理过程,发病率随年龄增长而上升,瓣膜间质细胞表型分化过程是瓣膜钙化的关键环节。脂质代谢异常作为瓣膜钙化的独立危险因素,在瓣膜局部形成损伤并诱导炎症反应,产生的炎症因子可以促进瓣膜间质细胞发生表型分化。非编码RNA可以从多方面调控炎症反应和成骨分化,影响瓣膜间质细胞表型分化进程,参与心脏瓣膜钙化的发生。